Porcine reproductive and respiratory syndrome (PRRS), originally known as “mystery swine disease,” was first recognized in the 1980's in the United States. The Lelystad virus (LV) strain of PRRSV was first isolated in Europe, (Wensvoot et al., “Mystery swine disease in the Netherlands: the isolation of Lelystad virus,” Vet Q 13, 121-130 (1991)) followed rapidly by the isolation of the North American prototype, strain VR-2332, in the United States (Collins et al. “Isolation of swine infertility and respiratory syndrome virus (Isolate ATCC-VR2332) in North America and experimental reproduction of the disease in gnotobiotic pigs,” J. Vet Diagn Invest 4, 117-126 (1992)). The principal clinical manifestation of the disease is characterized by a sudden onset of abortion and infertility in infected sows, and increased mortality rates in piglets which may result from secondary respiratory infections.
The etiological agent capable of reproducing the disease syndrome has been classified as a member of the Arteriviridae family of viruses and has been identified as an enveloped small spherical RNA virus. The virion particle has an average diameter of about 62 nm and a 25-30 nm nucleocapsid core surrounded by an envelope. The genome of PRRSV is a single-stranded, nonsegmented, 5′-capped and 3′-polyadenylated, positive-sense RNA of 15.1-15.5 kilobases in length, encoding at least seven demonstrated open reading frames (ORFs). The coding sequences are flanked by untranslated regions (UTR) at both the 5′ and 3′ ends of the genome. Viral structural proteins are expressed from six subgenomic mRNAs formed by a unique, yet unknown, discontinuous transcription mechanism by which the 5′ leader sequence noncontiguously joins with the body sequence located at the 3′ end of the viral genome. The viral replicase is encoded by ORFs 1a/1b, with the latter expressed via a −1 frameshift from the genomic RNA. ORFs 2 to 4 encode putative structural glycoproteins. ORF 5 encodes the major envelope glycoprotein of approximately 25 kDa. A non-glycosylated membrane protein of 18 kDa is encoded by ORF 6 and the 15 kDa nucleocapsid protein is encoded by ORF 7 (Snijder, E. J. and J. J., Meulenberg, “The molecular biology of arteriviruses,” J. Gen Virol 79, 961-979 (1998)).
The European and North American PRRSV genotypes are dramatically different both genetically and immunologically. Only about 45 percent nucleic acid identity and little serological cross-reaction between the genotypes have been reported. In general, European genotype PRRSV strains previously have been limited to Europe, while the viruses isolated from the rest of the world belong to the North American genotype, with ATCC VR-2332 as the prototype.
Recently, the swine industry has suffered from a severe form of PRRSV disease, characterized by increased abortion and mortality rates in pregnant sows. These atypical PRRSV outbreaks surprisingly have been reported in herds vaccinated with a commercially available live vaccine. These outbreaks clearly demonstrate a need for an improved vaccine to reduce or eliminate economic losses associated with swine morbidity and mortality caused by PRRSV disease. In addition, neurotropic PRRSV has been isolated from herds in which increased rates of abortion occurred, suggesting that PRRSV may be expanding its tissue tropism. The isolation of LV-like PRRSV in North America and the increased use of a vaccine derived from a North American strain in Europe are indications that the two genotypes of PRRSV are no longer geographically separated and are becoming endemic worldwide.
Full-length cDNA clones of positive-strand RNA viruses are important tools for the study of the biology of viruses (Boyer, J. C., “Infectious transcripts and cDNA clones of RNA viruses,” Virology 198, 415-426 (1994)). Infectious cDNA can be used to tackle fundamental questions regarding the mechanism of viral replication and pathogenesis. Moreover, full-length cDNA clones can be used to produce a genetically engineered molecular vaccine with desired genetic and immunologic traits. Although an infectious cDNA has been developed for the LV strain of PRRSV, there remains a need for a full-length North American PRRSV cDNA. Because of the expanding tropism and the emergence of the North American strain in herds worldwide, an infectious clone is needed to generate a more efficient molecular vaccine against PRRSV disease.